title: Efficient production of retroviruses using PLGA/bPEI-DNA nanoparticles and application for reprogramming somatic cells.
abstract: Reprogramming of somatic cells to pluripotent cells requires the introduction of factors driving fate switches. Viral delivery has been the most efficient method for generation of induced pluripotent stem cells. Transfection, which precedes virus production, is a commonly-used process for delivery of nucleic acids into cells. The aim of this study is to evaluate the efficiency of PLGA/ bPEI nanoparticles in transfection and virus production. Using a modified method of producing PLGA nanoparticles, PLGA/bPEI-DNA nanoparticles were examined for transfection efficiency and virus production yield in comparison with PLGA-DNA, bPEI-DNA nanoparticles or liposome-DNA complexes. After testing various ratios of PLGA, bPEI, and DNA, the ratio of 6:3:1 (PLGA:bPEI:DNA, w/w/w) was determined to be optimal, with acceptable cellular toxicity. PLGA/bPEI-DNA (6:3:1) nanoparticles showed superior transfection efficiency, especially in multiple gene transfection, and viral yield when compared with liposome-DNA complexes. The culture supernatants of HEK293FT cells transfected with PLGA/bPEI-DNA of viral constructs containing reprogramming factors (Oct4, Sox2, Klf4, or c-Myc) successfully and more efficiently generated induced pluripotent stem cell colonies from mouse embryonic fibroblasts. These results strongly suggest that PLGA/bPEI-DNA nanoparticles can provide significant advantages in studying the effect of multiple factor delivery such as in reprogramming or direct conversion of cell fate.
authors.affiliation: Medical Research Center for Ischemic Tissue Regeneration, Pusan National University School of Medicine, Yangsan, Republic of Korea ; Department of Physiology, Pusan National University School of Medicine, Yangsan, Republic of Korea.
authors.forename: Eun Jin
authors.initials: EJ
authors.lastname: Seo
authors.forename: Il Ho
authors.initials: IH
authors.lastname: Jang
authors.forename: Eun Kyoung
authors.initials: EK
authors.lastname: Do
authors.forename: Hyo Cheon
authors.initials: HC
authors.lastname: Cheon
authors.forename: Soon Chul
authors.initials: SC
authors.lastname: Heo
authors.forename: Yang Woo
authors.initials: YW
authors.lastname: Kwon
authors.forename: Geun Ok
authors.initials: GO
authors.lastname: Jeong
authors.forename: Ba Reun
authors.initials: BR
authors.lastname: Kim
authors.forename: Jae Ho
authors.initials: JH
authors.lastname: Kim
mesh_terms: D000818:Animals; D054337:Cell Dedifferentiation / Q000235:genetics*; D004247:DNA / Q000737:chemistry*; D005434:Flow Cytometry; D015316:Genetic Therapy / Q000379:methods; D057809:HEK293 Cells; D006801:Humans; D007150:Immunohistochemistry; D000090062:Kruppel-Like Factor 4; D019344:Lactic Acid / Q000737:chemistry*; D051379:Mice; D008856:Microscopy, Fluorescence; D041681:NIH 3T3 Cells; D053758:Nanoparticles / Q000737:chemistry*; D011094:Polyethyleneimine / Q000737:chemistry*; D011100:Polyglycolic Acid / Q000737:chemistry*; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D012190:Retroviridae / Q000235:genetics*; D014162:Transfection / Q000379:methods*
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid; D011094:Polyethyleneimine; D004247:DNA